Platelet activation in patients with peripheral vascular disease: reproducibility and comparability of platelet markers.

BACKGROUND: Many markers of platelet activation have been described but their reproducibility and comparability in patient populations are poorly defined. OBJECTIVES: We sought to compare markers of platelet and monocyte activation with platelet-monocyte aggregates, a proposed gold standard of in vivo platelet activation, and assess their reproducibility in patients with peripheral arterial disease: a population with substantial platelet activation, inflammation and risk of thrombotic events. PATIENTS/METHODS: Thirty patients with peripheral vascular disease attended on two occasions to permit within-day and between-day comparisons. In vivo platelet and monocyte activation were determined by flow-cytometric quantification of platelet-monocyte aggregation, platelet surface expression of P-selectin and CD40L, platelet-derived microparticles, and monocyte surface expression of CD40 and CD11b. Plasma concentrations of platelet-derived microparticles, soluble P-selectin and CD40L were measured by enzyme-linked immunosorbant assays. RESULTS: Platelet-monocyte aggregation (36.7±7.86%), and platelet surface expression of P-selectin (5.8±1.65%) and CD40L (3.3±1.45%) demonstrated comparable within-day (mean difference±co-efficient of reproducibility; 0.9±15.4%, 0.21±1.65% and 0.2±2.8% respectively) and between-day reproducibility (2.0±12.4%, 0.10±2.25% and 0.9±6.4% respectively). Platelet-monocyte aggregates correlated well with other platelet (r=0.30-0.50, P<0.02) and monocyte (r=0.27-0.47, P<0.03) activation markers. Flow cytometric and assay quantified platelet-derived microparticles showed poorer reproducibility (co-efficient of reproducibility >40). CONCLUSIONS: In patients with peripheral arterial disease, measurements of platelet-monocyte aggregates have good reproducibility and consistently reflect other markers of platelet and monocyte activation.

Vascular cell responsiveness to Toll-like receptor ligands in carotid atheroma.

BACKGROUND: Atherosclerosis is potentiated by stimulation of Toll-like receptors (TLRs), which serve to detect pathogen associated molecular patterns (PAMPs). However little is known of which PAMPs may be present in atheroma, or capable of stimulating inflammatory signalling in vascular cells. MATERIALS AND METHODS: DNA extracted from human carotid atheroma samples was amplified and sequenced using broad-range 16S gene specific primers to establish historical exposure to bacterial PAMPs. Responsiveness of primary human arterial and venous endothelial and smooth muscle cells to PAMPs specific for each of the TLRs was assessed by measurement of interleukin-8 secretion and E-selectin expression. RESULTS: Extracts of atheromatous tissue stimulated little or no signalling in TLR-transfected HEK-293 cells. However, sequencing of bacterial DNA amplified from carotid atheroma revealed the presence of DNA from 17 different bacterial genera, suggesting historical exposure to bacterial lipopeptide, lipopolysaccharide and flagellin. All cells examined were responsive to the ligands of TLR3 and TLR4, poly inosine:cytosine and lipopolysaccharide. Arterial cells were responsive to a wider range of PAMPs than venous cells, being additionally responsive to bacterial flagellin and unmethylated cytosine-phosphate-guanosine DNA motifs, the ligands of TLR5 and TLR9, respectively. Cells were generally unresponsive towards the ligands of human TLR7 and TLR8, loxoribine and single stranded RNA. Only coronary artery endothelial cells expressed TLR2 mRNA and responded to the TLR2 ligand Pam(3)CSK(4). CONCLUSIONS: Vascular cells are responsive to a relatively diverse range of TLR ligands and may be exposed, at least transiently, to ligands of TLR2, TLR4, TLR5 and TLR9 during the development of carotid atheroma.

Harnessing the preconditioning phenomenon: does remote organ ischaemia provide the answer?

Despite progress in defining the cellular mechanisms of the ischaemic preconditioning phenomenon, its conversion into convenient clinical practice has been slow. The possibility that an innate mechanism of tissue resistance to ischaemia could be harnessed as a clinical tool is an attractive and enticing prospect.

Comparison between the Datex-Ohmeda M-COVX metabolic monitor and the Deltatrac II in mechanically ventilated patients.

OBJECTIVE: To compare the M-COVX and the Deltatrac II metabolic monitors under clinical conditions. DESIGN: Prospective clinical comparison. SETTING: A general Intensive Care Unit of a university hospital. PATIENTS: Twenty mechanically ventilated critically ill patients. INTERVENTIONS: The monitors were compared at FiO(2) 0.3, 0.5, and 0.7 in each patient where possible. MEASUREMENTS AND RESULTS: Pulmonary gas exchange measurements were recorded using the two monitors sequentially (Deltatrac(before), M-COVX, Deltatrac(after)). Each measurement consisted of five consecutive 1-min readings of VO(2) and VCO(2). We compared the Deltatrac(before) with the Deltatrac(after) and the mean of the Deltatrac with the M-COVX. There was no clinically significant bias between the two monitors for VO(2) or VCO(2) but the limits of agreement (LOA) were wide (bias +/-95% LOA: VCO(2) -13 +/- 30 ml/min, -8 +/- 36 ml/min, 7 +/- 50 ml/min; VO(2) -7 +/- 50 ml/min, -5 +/- 56 ml/min, 6 +/- 64 ml/min, at FiO(2) 0.3, 0.5, and 0.7, respectively). The Deltatrac before and after measurements displayed good agreement for VCO(2) but poorer agreement for VO(2) (bias +/- 95% LOA: VCO(2) 0 +/- 18 ml/min, -6 +/- 16 ml/min, -1 +/- 12 ml/min; VO(2) 2 +/- 12 ml/min, 3 +/- 38 ml/min, 10 +/- 42 ml/min, at FiO(2) 0.3, 0.5, and 0.7, respectively). Using within-patient standard deviation as a measure of reproducibility suggested that for VO(2) the M-COVX performed better than the Deltatrac at high FiO(2), and for VCO(2) Deltatrac was better at lower FiO(2). CONCLUSIONS: The M-COVX is a suitable integrated device for measuring metabolic gas exchange in ventilated patients.